Abstract
BACKGROUND: Esophageal squamous cell carcinoma (ESCC) remains a lethal malignancy with limited therapeutic options. JMJD2A (KDM4A) promotes tumor progression, epithelial–mesenchymal transition, and immune evasion, and it has been implicated in ferroptosis resistance. Ultrasound-targeted microbubble destruction (UTMD) can enhance local delivery of nucleic acids. We hypothesized that UTMD-mediated delivery of siRNA against JMJD2A (si-JMJD2A) would trigger ferroptosis, curb metastatic traits, remodel the tumor immune microenvironment, and improve responsiveness to anti-PD-L1 therapy. METHODS: si-JMJD2A was formulated with UTMD carriers and characterized for size and surface charge. ESCC cell lines (K150 and K450) were used to assess transfection efficiency, JMJD2A knockdown, proliferation and colony formation, migration and invasion, and ferroptosis readouts, including lipid reactive oxygen species and rescue with ferroptosis inhibitors. Macrophage polarization and chemotaxis were profiled to evaluate tumor-induced immune remodeling. In vivo efficacy was tested in mouse models of ESCC, with end points including primary tumor growth, pulmonary metastasis, survival, and combination therapy with anti-PD-L1. RESULTS: UTMD enhanced siRNA uptake and produced robust JMJD2A silencing in ESCC cells. Functionally, si-JMJD2A delivered by UTMD suppressed proliferation, clonogenicity, migration, and invasion while inducing hallmarks of ferroptosis; these effects were mitigated by ferroptosis antagonism, supporting on-pathway activity. UTMD-si-JMJD2A reduced M2-like polarization and macrophage chemotaxis, consistent with relief of tumor-driven immune suppression. In mice, UTMD-mediated si-JMJD2A inhibited tumor growth and metastasis and improved survival. Combining UTMD-si-JMJD2A with anti-PD-L1 further enhanced antitumor efficacy compared with monotherapy. CONCLUSIONS: This study establishes image-guided UTMD as a nonviral, localized delivery strategy for therapeutic JMJD2A silencing in ESCC. UTMD-mediated si-JMJD2A delivery induces ferroptosis, suppresses metastatic traits, and enhances the antitumor efficacy of PD-L1 blockade, supporting UTMD-enabled gene silencing as a translational approach for ESCC. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12672-026-04583-3.